Identification of locally isolated entomopathogenic Fusarium species from the soil of Changa Manga Forest, Pakistan and evaluation of their larvicidal efficacy against Aedes aegypti

Dengue fever vectored by the mosquito Aedes aegypti is one of the most rapidly spreading insect-borne diseases. Current reliance of dengue vector control is mostly on chemical insecticides. Growing insecticide resistance in the primary mosquito vector, Aedes aegypti, limits the effectiveness of vector control through chemical insecticides. These chemical insecticides also have negative environmental impacts on animals, plants and human health. Myco-biocontrol agents are naturally occurring organisms and are found to be less damaging to the environment as compared to chemical insecticides. In the present study, entomopathogenic potential of local strains of fungi isolated from soil was assessed for the control of dengue vector. Local fungal isolates presents better alternative to introducing a foreign biocontrol strain, as they may be better adapted to environmental conditions of the area to survive and may have more entomopathogenic efficacy against target organism. Larvicidal efficacy of Fusarium equiseti and Fusarium proliferatum was evaluated against Aedes aegypti. Local strains of F. equiseti (MK371718) and F. proliferatum (MK371715) were isolated from the soil of Changa Manga Forest, Pakistan by using insect bait method. Larvicidal activity of two Fusarium spp. was tested against forth instar larvae of A. aegypti in the laboratory, using concentrations 105, 106, 107 and 108 conidia /ml. LC50 values for F. equiseti after 24h, 48h, 72h and 96h of exposure were recorded as 3.8x 108, 2.9x107, 2.0x107, and 7.1x106 conidia /ml respectively while LC50 values for F. proliferatum were recorded as 1.21x108, 9.6x107, 4.2x107, 2.6x107 conidia /ml respectively after 24h, 48h, 72h and 96h of exposure. The results indicate that among two fungal strains F. equiseti was found to be more effective in terms of its larvicidal activity than F. proliferatum against larvae of A. aegypti.

Standard selection treatments with sulfadiazine limit Plasmodium yoelii host-to-vector transmission

Some early antimalarial drugs have been repurposed for experimental applications, thus extending their utility well beyond the point when resistance becomes prevalent in circulating parasite populations. One such drug is sulfadiazine, which is an analog of p-aminobenzoic acid (pABA), and acts as a competitive inhibitor of dihydropteroate synthase, which is an essential enzyme in the parasite's folate synthesis pathway that is required for DNA synthesis. Sulfadiazine treatment of mice infected with P. yoelii and P. berghei is routinely used to enrich for gametocytes by killing asexual blood stage parasites, but it is not well known if the exposed gametocytes are perturbed or if there is a detrimental effect on transmission. To determine if there was a significant effect of sulfadiazine exposure upon host-to-vector transmission, we transmitted Plasmodium yoelii (17XNL strain) parasites to Anopheles stephensi mosquitoes and evaluated the prevalence of infection (percent of mosquitoes infected) and intensity of infection (number of oocysts per infected mosquito) under different sulfadiazine treatment conditions of the mouse or of the mosquitoes. We observed that parasites exposed to sulfadiazine either in the mouse host or in the mosquito vector had a reduction in both the number of mosquitoes that became infected and in the intensity of infection compared to untreated controls. We also observed that provision of freshly prepared pABA in the mosquito sugar water could only marginally overcome the defects caused by sulfadiazine treatment. In contrast, we determined that gametocytes exposed to sulfadiazine were able to be fertilized and develop into morphologically mature ookinetes in vitro, and thus that sulfadiazine exposure in the host may be reversible if the drug is washed out and the parasites are supplemented with pABA in the culture media. Overall, this indicates that sulfadiazine dampens host-to-vector transmission, and that this inhibition can only be partially overcome by exposure to fresh pABA in vivo and in vitro. Because gametocytes are of great interest for developing transmission blocking interventions, we recommend that less disruptive approaches for gametocyte enrichment be used in order to study minimally perturbed parasites.

Integrated control of Aedes albopictus in Southwest Germany supported by the Sterile Insect Technique

Background The invasive species Aedes albopictus, commonly known as the Asian tiger mosquito, has undergone extreme range expansion by means of steady introductions as blind passengers in vehicles traveling from the Mediterranean to south-west Germany. The more than 25 established populations in the State of Baden-Württemberg, Palatine and Hesse (south-west Germany) have become a major nuisance and public health threat. Aedes albopictus deserves special attention as a vector of arboviruses, including dengue, chikungunya and Zika viruses. In Germany, Ae. albopictus control programs are implemented by local communities under the auspices of health departments and regulatory offices. Methods The control strategy comprised three pillars: (i) community participation (CP) based on the elimination of breeding sites or improved environmental sanitation, using fizzy tablets based on Bacillus thuringiensis israelensis (fizzy Bti tablets; Culinex® Tab plus); (ii) door-to-door (DtD) control by trained staff through the application of high doses of a water-dispersible Bti granular formulation (Vectobac® WG) aimed at achieving a long-lasting killing effect; and (iii) implementation of the sterile insect technique (SIT) to eliminate remaining Ae. albopictus populations. Prior to initiating large-scale city-wide treatments on a routine basis, the efficacy of the three elements was evaluated in laboratory and semi-field trials. Special emphasis was given to the mass release of sterile Ae. albopictus males. Results More than 60% of the local residents actively participated in the first pillar (CP) of the large-scale control program. The most effective element of the program was found to be the DtD intervention, including the application of Vectobac® WG (3000 ITU/mg) to potential breeding sites (10 g per rainwater container, maximum of 200 l = maximum of approx. 150,000 ITU/l, and 2.5 g per container < 50 l) with a persistence of at least 3 weeks. In Ludwigshafen, larval source management resulted in a Container Index for Ae. albopictus of < 1% in 2020 compared to 10.9% in 2019. The mean number of Aedes eggs per ovitrap per 2 weeks was 4.4 in Ludwigshafen, 18.2 in Metzgergrün (Freiburg) (SIT area) and 22.4 in the control area in Gartenstadt (Freiburg). The strong reduction of the Ae. albopictus population by Bti application was followed by weekly releases of 1013 (Ludwigshafen) and 2320 (Freiburg) sterile Ae. albopictus males per hectare from May until October, resulting in a high percentage of sterile eggs. In the trial areas of Ludwigshafen and Frieburg, egg sterility reached 84.7 ± 12.5% and 62.7 ± 25.8%, respectively; in comparison, the natural sterility in the control area was 14.6 ± 7.3%. The field results were in line with data obtained in cage tests under laboratory conditions where sterility rates were 87.5 ± 9.2% after wild females mated with sterile males; in comparison, the sterility of eggs laid by females mated with unirradiated males was only 3.3 ± 2.8%. The overall egg sterility of about 84% in Ludwigshafen indicates that our goal to almost eradicate the Ae. albopictus population could be achieved. The time for inspection and treatment of a single property ranged from 19 to 26 min depending on the experience of the team and costs 6–8 euros per property. Conclusions It is shown that an integrated control program based on a strict monitoring scheme can be most effective when it comprises three components, namely CP, DtD intervention that includes long-lasting Bti-larviciding to strongly reduce Ae. albopictus populations and SIT to reduce the remaining Ae. albopictus population to a minimum or even to eradicate it. The combined use of Bti and SIT is the most effective and selective tool against Ae. albopictus, one of the most dangerous mosquito vector species. Graphical Abstract

Aedes aegypti abundance, larval indices and risk for dengue virus transmission in Kinondoni district, Tanzania

Background Tanzania has experienced periodic dengue outbreaks with increased incidence since 2010. However, there is limited information on vector dynamics and transmission risk in most parts of the country. This study was conducted to determine Aedes mosquito abundance, larval indices and dengue virus infection rate as risk indicators for DENV transmission in Kinondoni district, Dar es Salaam, Tanzania. Methods A cross-sectional study was conducted in three wards of Kinondoni district in Tanzania between December 2019 and January 2020. In each ward, three streets were randomly selected for adult and immature mosquito sampling. The adult mosquitoes were collected using Mosquito Magnet traps, while mosquito larvae and pupae were inspected in water-holding containers in the selected household compounds. The detection of dengue virus (DENV) in female Aedes mosquitoes was done using a one-step reverse transcription–polymerase chain reaction (RT–PCR) method. Results Of the 1416 adult female mosquitoes collected, Ae. aegypti accounted for 16.8% (n = 238). A total of 333 water-holding containers were inspected and 201 (60.4%) had at least an Aedes larvae or pupae. Water-holding containers supporting the breeding of Aedes larvae and pupae included discarded car tires, flowerpots and small and large plastic containers. The overall House Index, Container Index and Breteau Index were 55.1%, 60.4% and 114.2, respectively. None of the 763 female Aedes mosquitoes tested by RT–PCR was found to be infected with DENV. Conclusion The presence and abundance Ae. aegypti mosquitoes and the large proportion of water-holding containers infested with the mosquito larvae and pupae put residents of Kinondoni district at high risk of DENV transmission. Our findings emphasize the need for continuous mosquito vector surveillance and control to prevent the possibility of future DENV outbreaks in Tanzania.

Antitrypanosomal Effect of Hydromethanolic Extract of Solanum anguivi Lam on Field Isolates of Trypanosoma congolense Infected Mice

Introduction. Trypanosomiasis is one of the world’s most serious infectious diseases caused by Trypanosoma parasites. Concern about resistance to conventional antitrypanosomal drugs, mosquito vector resistance to existing insecticide side effects of existing antitrypanosomal drugs justifies the urgent need for more effective, tolerable, and affordable drugs. Objective. The present study is aimed at determining the in vivo antitrypanosomal effect of the hydromethanolic extracts of Solanum anguivi fruit extracts against the field isolates of T. congolense. Methods. The 80% methanol extracts of S. anguivi fruits were prepared by cold maceration technique. In vivo curative tests were done to check the effect of plant extract against T. congolense in Swiss albino mice. Plant extracts were administered at doses of 100, 200, and 400 mg/kg/body weight. Acute toxicity of the extracts at 2000 mg/kg was performed according to OECD guidelines. Data obtained from the experiment were analyzed using one-way ANOVA followed by Tukey test. Results. This study indicated that extract did not exhibit any sign of acute toxicity up to 2000 mg/kg/body weight. In curative test, extracts reduced parasitemia, preventing the drop in packed cell volume and body weight significantly ( p < 0.05 ), compared to control. Groups provided with the extract before infection got prolonged incubation period with chemoprophylactic effect at the doses of 100, 200, and 400 mg/kg. Phytochemical analysis showed presence of flavonoids, steroids, triterpens, saponins, glycosides, tannins, and alkaloids. Conclusion. The extract showed promising curative. Further effort is required to isolate and purify specific compounds responsible for antitrypanosomal activity of studied plant.

Potensi Culex quinquefasciatus sebagai Vektor Filariasis dan Kondisi Lingkungan di Kota Pekalongan

Pekalongan City was still an endemic area of Lymphatic Filariasis (LF). Twice cycles ofMass Drug Administration (MDA) had been implemented, but the Microfi laria rate (Mf rate) wasstill more than 1%. This observational research aimed to study the potential of Culex quinquefasciatusas a vector of LF in Pekalongan City. A Cross-sectional design was chosen as an approach in compilinginformation related to environmental factors and mosquitoes. The population of this research washouses located in Jenggot and Kertoharjo village with thirty-one houses, located around fi lariasiscases, purposively selected as samples. Variables studied in this research were environmental factorsand mosquito vectors. Data collection using observation, and laboratory examination throughmosquito dissection. Data would be analyzed descriptively. This study found that there were 8 Culexquinquefasciatus tested positive L3 fi larial worm (infective rate 4.39%). There were 74.2% of houseshad mosquitoes’ breeding sites around them. The breeding sites were found at domestic waste disposal,drainage in front of the house, and infi ltration well for liquid waste. As much as 86.2% of the breedingsites contained mosquito larvae. This research concluded that Cx. quinquefasciatus was confi rmedpositive fi laria worm thus establish as mosquito vector for Lymphatic Filariasis in Pekalongan City.The breeding sites related to the mosquito development were small water bodies, drainage in front ofand around the house.

Species Composition, Abundance and Population Structure of Malaria Vectors in Two Villages of Sudan

BackgroundFrequent monitoring of mosquito vector population is a strategy of great importance for reducing risks of disease occurrence. In Sudan, malaria is still a big threat to public health. Insecticide-based control has been undertaken for years, but there is no noticeable decrease of malaria infection nationwide.ObjectiveTo overcome this situation, a better understanding of the breeding ecology of the vectors is relevant. Here, we investigate the species composition of malaria vectors, seasonal abundance and population structure in two different villages.MethodologyMonthly samplings were performed in Abu Algoni (Sennar State) and Algerif West (Khartoum State) from June 2010 to May 2011. During each visit, immature stages were collected from potential breeding sites using dipping technique. In addition, adults were collected indoors from houses by aspiration and indoor pyrethrum spray methods. Mosquitoes were identified morphologically, the Anopheles gambiae complex diagnosed using PCR and the physiological status of females determined based on appropriate techniques. Environmental parameters namely temperature, rainfall and humidity were measured.ResultsA total of 4,932 mosquitoes comprising of 3047 larvae and 1885 adults (males and females combined) were sampled. Of these, 88.9% were collected from Abu Algoni while 11.1% were from Algerif West. Two species, An. rufipes and An. arabiensis were encountered. Anopheles rufipes was only found in Abu Algoni, while the latter was found in both villages, where it represented more than 99% of the total collection.Mosquitoes were found breeding in many types of places including canals, temporary pools of water, animal hooves, water from broken pipes, and water storage containers. No significant correlation was found between female and temperature (p>0.05). Significant correlation difference was observed between number of females of An. arabiensis and rainfall (p<0.05) and humidity (p<0.01).ConclusionsAnopheles arabiensis is the only member of An. gambiae complex detected in the present study. Seasonal abundance of An. arabiensis was observed with most during the wet rainy season in both villages. This could be associated with the availability of more breeding sites created by the rainfall. The majority were parous which indicates high survival rates and thus high vectorial capacity in transmitting malaria.

Habitat Segregation Patterns of Container Breeding Mosquitos: The Role of Urban Heat Islands, Vegetation Cover, and Income Disparity in Cemeteries of New Orleans

Aedes aegypti and Aedes albopictus are important pathogen-carrying vectors that broadly exhibit similar habitat suitability, but that differ at fine spatial scales in terms of competitive advantage and tolerance to urban driven environmental parameters. This study evaluated how spatial and temporal patterns drive the assemblages of these competing species in cemeteries of New Orleans, LA, applying indicators of climatic variability, vegetation, and heat that may drive habitat selection at multiple scales. We found that Ae. aegypti was well predicted by urban heat islands (UHI) at the cemetery scale and by canopy cover directly above the cemetery vase. As predicted, UHI positively correlate to Ae. aegypti, but contrary to predictions, Ae. aegypti, was more often found under the canopy of trees in high heat cemeteries. Ae. albopictus was most often found in low heat cemeteries, but this relationship was not statistically significant, and their overall abundances in the city were lower than Ae. aegypti. Culex quinquefasciatus, another important disease vector, was also an abundant mosquito species during the sampling year, but we found that it was temporally segregated from Aedes species, showing a negative association to the climatic variables of maximum and minimum temperature, and these factors positively correlated to its more direct competitor Ae. albopictus. These findings help us understand the mechanism by which these three important vectors segregate both spatially and temporally across the city. Our study found that UHI at the cemetery scale was highly predictive of Ae. aegypti and strongly correlated to income level, with low-income cemeteries having higher UHI levels. Therefore, the effect of excessive heat, and the proliferation of the highly competent mosquito vector, Ae. aegypti, may represent an unequal disease burden for low-income neighborhoods of New Orleans that should be explored further. Our study highlights the importance of considering socioeconomic aspects as indirectly shaping spatial segregation dynamics of urban mosquito species.

Integrated Control of Aedes Aegypti With Parasitic Nematodes, Biological Control Agents, Chemical Insecticides and Plant Essential Oil

Aedes aegypti can transmit dengue fever, yellow fever, Chikungunya fever, Zika virus disease and vector density control is the most effective way to prevent these infectious diseases. However, the extensive use of chemical pesticides has caused a series of problems, such as environmental pollution, killing non-target organisms and so on. In this study, a parasitic nematode, Romanomermis wuchangensis was used in the larviciding evaluation of Ae. aegypti, while the activity of four chemical insecticides and biological control agents were tested. Besides, Mentha haplocalyx essential oil was isolated and its olfactory physiological function with OBP1 protein of Ae. aegypti antenna was measured by the prokaryotic expression and fluorescence competitive binding assay. Compared with the control group, R. wuchangensis indicated high efficiency and environmental friendliness in the control of Ae. aegypti. After the second instar larvae were parasitized, the mortality of two treatment groups exceeded 75%. Compared to control group, the quantitative real-time PCR analysis results demonstrated that SOD, POD and CAT genes had obvious high expression levels in the nematodes parasitic groups. The antioxidant enzyme test results also exhibited obvious difference of SOD, CAT and POD during the nematode parasitic period. Besides, Bacillus thuringiensis (Bti) and chemical insecticide experimental results also showed great insecticidal efficacy against mosquito larvae. Five chemical components including Menthol, Pinene, Limonene, Isopulegol and Pulegone were identified from M. haplocalyx and exhibited great binding ability with AaegOBP1 protein. Present results illustrated that the integrated application of these various mosquito vector control methods in the future has broad prospects.

A transcriptomic atlas of Aedes aegypti reveals detailed functional organization of major body parts and gut regional specializations in sugar-fed and blood-fed adult females

Mosquito vectors transmit numerous pathogens, but large gaps remain in our understanding of their physiology. To facilitate future explorations of mosquito biology, with specific attention to the major vector Aedes aegypti, we have created Aegypti-Atlas (http://aegyptiatlas.buchonlab.com/), an online resource hosting RNAseq profiles of Ae. aegypti body parts (head, thorax, abdomen, gut, Malpighian tubules, and ovaries), gut regions (crop, proventriculus, anterior and posterior midgut, and hindgut), and a time course of blood meal digestion in the gut. Using Aegypti-Atlas, we provide new insights into the regionalization of gut function, blood feeding response, and immune defenses. We find that the anterior and posterior regions of the mosquito midgut possess clearly delineated digestive specializations which are preserved in the blood-fed state. Blood feeding initiates the sequential transcriptional induction and repression/depletion of multiple cohorts of peptidases throughout blood meal digestion. With respect to defense, immune signaling components, but not recognition or effector molecules, show enrichment in ovaries. Basal expression of antimicrobial peptides is dominated by two genes, holotricin and gambicin, that are expressed in the carcass and the digestive tissues, respectively, in a near mutually exclusive manner. In the midgut, gambicin and other immune effector genes are almost exclusively expressed in the anterior regions, while the posterior midgut exhibits the hallmarks of immune tolerance. Finally, in a cross-species comparison between the midguts of Ae. aegypti and Anopheles gambiae, we observe that regional digestive and immune specializations are closely conserved, indicating that our data may yield inferences that are broadly relevant to multiple mosquito vector species. We further demonstrate that the expression of orthologous genes is highly correlated, with the exception of a ‘species signature’ comprising a small number of highly/disparately expressed genes. With this work, we show the potential of Aegypti-Atlas to unlock a more complete understanding of mosquito biology.